Emulsion vehicle for poorly soluble drugs

ABSTRACT

An emulsion of α-tocopherol, stabilized by biocompatible surfactants, as a vehicle or carrier for therapeutic drugs, which is substantially ethanol free and which can be administered to animals or humans various routes is disclosed. Also included in the emulsion is PEGylated vitamin E. PEGylated α-tocopherol includes polyethylene glycol subunits attached by a succinic acid diester at the ring hydroxyl of vitamin E and serves as a primary surfactant, stabilizer and a secondary solvent in emulsions of α-tocopherol.

RELATED APPLICATIONS

This application is a non-provisional application based on U.S.Provisional Application No. 60/034,188 filed Jan. 7, 1997 and U.S.Provisional Application No. 60/048,840 filed Jun. 6, 1997 and claims thebenefit of these filing dates under 35 U.S.C. § 119(e) for prioritypurposes.

BACKGROUND OF THE INVENTION

Hundreds of medically useful compounds are discovered each year, butclinical use of these drugs is possible only if a drug delivery vehicleis developed to transport them to their therapeutic target in the humanbody. This problem is particularly critical for drugs requiringintravenous injection in order to reach their therapeutic target ordosage but which are water insoluble or poorly water soluble. For suchhydrophobic compounds, direct injection may be impossible or highlydangerous, and can result in hemolysis, phlebitis, hypersensitivity,organ failure and/or death. Such compounds are termed by pharmacists“lipophilic”, “hydrophobic”, or in their most difficult form,“amphiphobic”.

A few examples of therapeutic substances in these categories areibuprofen, diazepam, griseofulvin, cyclosporin, cortisone, proleukin,etoposide and paclitaxel. Kagkadis, K A et al. (1996) PDA J Pharm SciTech 50(5):317-323; Dardel, O. 1976. Anaesth Scand 20:221-24. Sweetana,S and M J U Akers. (1996) PDA J Pharm Sci Tech 50(5):330-342.

Administration of chemotherapeutic or anti-cancer agents is particularlyproblematic. Low solubility anti-cancer agents are difficult tosolubilize and supply at therapeutically useful levels. On the otherhand, water-soluble anti-cancer agents are generally taken up by bothcancer and non-cancer cells thereby exhibiting non-specificity.

Efforts to improve water-solubility and comfort of administration ofsuch agents have not solved, and may have worsened, the two fundamentalproblems of cancer chemotherapy: 1) non-specific toxicity and 2) rapidclearance form the bloodstream by non-specific mechanisms. In the caseof cytotoxins, which form the majority of currently availablechemotherapies, these two problems are clearly related. Whenever thetherapeutic is taken up by noncancerous cells, a diminished amount ofthe drug remains available to treat the cancer, and more importantly,the normal cell ingesting the drug is killed.

To be effective in treating cancer, the chemotherapeutic must be presentthroughout the affected tissue(s) at high concentration for a sustainedperiod of time so that it may be taken up by the cancer cells, but notat so high a concentration that normal cells are injured beyond repair.Obviously, water soluble molecules can be administered in this way, butonly by slow, continuous infusion and monitoring, aspects which entailgreat difficulty, expense and inconvenience.

A more effective method of administering a cancer therapeutic,particularly a cytotoxin, is in the form of a dispersion of oil in whichthe drug is dissolved. These oily particles are made electricallyneutral and coated in such a way that they do not interact with plasmaproteins and are not trapped by the reticuloendothelial system (RES),instead remaining intact in the tissue or blood for hours, days or evenweeks. In most cases, it is desirable if the particles also distributethemselves into the surrounding lymph nodes which are injected at thesite of a cancer. Nakamoto, Y et al. (1975) Chem Pharm Bull23(10):2232-2238. Takahashi, T et al. (1977) Tohoku J Exp Med123:235-246. In many cases direct injection into blood is the route ofchoice for administration. Even more preferable, following intravenousinjection, the blood-borne particles may be preferentially captured andingested by the cancer cells themselves. An added advantage of aparticulate emulsion for the delivery of a chemotherapeutic is thewidespread property of surfactants used in emulsions to overcomemultidrug resistance.

For drugs that cannot be formulated as an aqueous solution, emulsionshave typically been most cost-effective and gentle to administer,although there have been serious problems with making them sterile andendotoxin free so that they may be administered by intravenousinjection. The oils typically used for pharmaceutical emulsions includesaponifiable oils from the family of triglycerides, for example, soybeanoil, sesame seed oil, cottonseed oil, safflower oil and the like.Hansrani, P K et al., (1983) J Parenter Sci Technol 37:145-150. One ormore surfactants are used to stabilize the emulsion, and excipients areadded to render the emulsion more biocompatible, stable and less toxic.Lecithin from egg yolks or soybeans is a commonly used surfactant.Sterile manufacturing can be accomplished by absolute sterilization ofall the components before manufacture, followed by absolutely aseptictechnique in all stages of manufacture. However, improved ease ofmanufacture and assurance of sterility is obtained by terminalsterilization following sanitary manufacture, either by heat or byfiltration. Unfortunately, not all emulsions are suitable for heat orfiltration treatments.

Stability has been shown to be influenced by the size and homogeneity ofthe emulsion. The preferred emulsion consists of a suspension ofsub-micron particles, with a mean size of no greater than 200nanometers. A stable dispersion in this size range is not easilyachieved, but has the benefit that it is expected to circulate longer inthe bloodstream. Further, less of the stable dispersion is phagocytizednon-specifically by the reticuloendothelial system. As a result the drugis more likely to reach its therapeutic target. Thus, a preferred drugemulsion will be designed to be actively taken up by the target cell ororgan, and is targeted away from the RES.

The use of vitamin E in emulsions is known. In addition to the hundredsof examples where vitamin E in small quantities [for example, less than1%, R T Lyons. Pharm Res 13(9): S-226, (1996) “Formulation developmentof an injectable oil-in-water emulsion containing the lipophilicantioxidants K-tocopherol and P-carotene”] is used as an anti-oxidant inemulsions, the first primitive, injectable vitamin E emulsions per sewere made by Hidiroglou for dietary supplementation in sheep and forresearch on the pharmacokinetics of vitamin E and its derivatives.Hidiroglou M and Karpinski K. (1988) Brit J Nutrit 59:509-518.

For mice, an injectable form of vitamin E was prepared by Kato andcoworkers. Kato Y., et al. (1993) Chem Pharm Bull 41(3):599-604.Micellar solutions were formulated with Tween 80, Brij 58 and HCO-60.Isopropanol was used as a co-solvent, and was then removed by vacuumevaporation; the residual oil glass was then taken up in water withvortexing as a micellar suspension. An emulsion was also prepared bydissolving vitamin E with soy phosphatidycholine (lecithin) and soybeanoil. Water was added and the emulsion prepared with sonication.

In 1983, E-Ferol, a vitamin E emulsion was introduced for vitamin Esupplementation and therapy in neonates. Alade S L et al. (1986)Pediatrics 77(4):593-597. Within a few months over 30 babies had died asa result of receiving the product, and the product was promptlywithdrawn by FDA order. The surfactant mixture used in E-Ferol toemulsify 25 mg/mL vitamin E consisted of 9% Tween 80 and 1% Tween 20.These surfactants seem ultimately to have been responsible for theunfortunate deaths. This experience illustrates the need for improvedformulations and the importance of selecting suitable biocompatiblesurfactants and carefully monitoring their levels in parenteralemulsions and.

An alternative means of solubilizing low solubility compounds is directsolubilization in a non-aqueous milieu, for example alcohol (such asethanol) dimethylsulfoxide or triacetin. An example in PCT applicationWO 95/11039 describes the use of vitamin E and the vitamin E derivativeTPGS in combination with ethanol and the immuno-suppressant moleculecyclosporin. Alcohol-containing solutions can be administered with care,but are typically given by intravenous drip to avoid the pain, vascularirritation and toxicity associated with bolus injection of thesesolutions.

Problems with pharmaceutical formulations in non-aqueous solvents andsolubilizers such as alcohol (ethanol, isopropanol, benzyl alcohol,etc.) relate to the ability of these solvents to extract toxicsubstances, for example plasticizers, from their containers. The currentcommercial formulation for the anti-cancer drug paclitaxel, for example,consists of a mixture of hydroxylated castor oil and ethanol, andrapidly extracts plasticizers such as di-(2-ethylhexyl)-phthalate fromcommonly used intravenous infusion tubing and bags. Adverse reactions tothe plasticizers have been reported, such as respiratory distress,necessitating the use of special infusion systems at extra expense andtime. Waugh, et al. (1991) Am J Hosp Pharmacists 48:1520.

In light of these problems, it can be seen that the ideal emulsionvehicle would be inexpensive, non-irritating or even nutritive andpalliative in itself, terminally sterilizable by either heat orfiltration, stable for at least 1 year under controlled storageconditions, accommodate a wide variety of water insoluble and poorlysoluble drugs and be substantially ethanol-free. In addition to thosedrugs which are lipophilic and dissolve in oils, also needed is avehicle which will stabilize, and carry in the form of an emulsion,drugs which are poorly soluble in lipids and in water.

SUMMARY OF THE INVENTION

In order to meet these needs, the present invention is directed topharmaceutical compositions including: α-tocopherol, a surfactant ormixtures of surfactants, with and without an aqueous phase, and atherapeutic agent wherein the composition is in the form of an emulsion,micellar solution or a self-emulsifying drug delivery system. In apreferred form, the solution is substantially ethanol-free.

The pharmaceutical compositions can be stabilized by the addition ofvarious amphiphilic molecules, including anionic, nonionic, cationic,and zwitterionic surfactants. Preferably, these molecules are PEGylatedsurfactants and optimally PEGylated α-tocopherol.

The amphiphilic molecules further include surfactants such as ascorbyl-6palmitate; stearylamine; sucrose fatty acid esters, various vitamin Ederivatives and fluorine-containing surfactants, such as the Zonyl brandseries and a polyoxypropylene-polyoxyethylene glycol nonionic blockcopolymer.

The therapeutic agent of the emulsion may be a chemotherapeutic agentpreferably a taxoid analog and most preferably, paclitaxel.

The emulsions of the invention can comprise an aqueous medium when inthe form of an emulsion or micellar solution. This medium can containvarious additives to assist in stabilizing the emulsion or in renderingthe formulation biocompatible.

The pharmaceutical compositions of the invention are typically formed bydissolving a therapeutic agent in ethanol to form a therapeutic agentsolution. α-tocopherol is then added to the therapeutic agent solutionto form an α-tocopherol and therapeutic agent solution. Next, theethanol is removed to form a substantially ethanol-free α-tocopherol andtherapeutic agent solution. The substantially ethanol free α-tocopheroland therapeutic agent solution is blended with and without an aqueousphase incorporating a surfactant to form a pre-emulsion. For IV deliverythe pre-emulsion is then homogenized to form a fine emulsion. For oraldelivery, the pre-emulsion is typically encapsulated in a gelatincapsule.

BRIEF DESCRIPTION OF THE DRAWINGS

The invention will be better understood by reference to the figures, inwhich:

FIG. 1A shows the particle size of a paclitaxel emulsion (QWA) at 7° C.over time;

FIG. 1B shows the particle size of a paclitaxel emulsion (QWA) at 25° C.over time;

FIG. 2 is an HPLC chromatogram showing the integrity of a paclitaxel inan emulsion as described in Example 5;

FIG. 3A shows the paclitaxel concentration of a paclitaxel emulsion(QWA) at 4° C. over time;

FIG. 3B shows the paclitaxel concentration of a paclitaxel emulsion(QWA) at 25° C. over time; and

FIG. 4 shows the percentage of paclitaxel released over time from threedifferent emulsions. The symbol  represents the percentage ofpaclitaxel released over time from an emulsion commercially availablefrom Bristol Myers Squibb. The symbol ▾ represents the percentage ofpaclitaxel released over time from an emulsion of this inventioncontaining 6 mg/ml paclitaxel (QWA) as described in Example 6. Thesymbol ⋄ represents the percentage of paclitaxel released over time froman emulsion of this invention (QWB) containing 7 mg/ml paclitaxel asdescribed in Example 7.

DETAILED DESCRIPTION OF THE INVENTION

To ensure a complete understanding of the invention the followingdefinitions are provided:

α-tocopherol: α-tocopherol, also known as vitamin E, is an organicmolecule with the following chemical structure (Scheme I):

In addition to its use as a primary solvent, α-tocopherol and itsderivatives are useful as a therapeutic agents.

Surfactants: Surface active group of amphiphilic molecules which aremanufactured by chemical processes or purified from natural sources orprocesses. These can be anionic, cationic, nonionic, and zwitterionic.Typical surfactants are described in Emulsions: Theory and Practice,Paul Becher, Robert E. Krieger Publishing, Malabar, Fla., 1965;Pharmaceutical Dosage Forms: Dispersed Systems Vol. I, Martin M. Rigear,Surfactants and U.S. Pat. No. 5,595,723 which is assigned to theassignee of this invention, Sonus Pharmaceuticals. All of thesereferences are hereby incorporated by reference.

TPGS: TPGS or PEGylated vitamin E is a vitamin E derivative in whichpolyethylene glycol subunits are attached by a succinic acid diester atthe ring hydroxyl of the vitamin E molecule. TPGS stands forD-α-tocopherol polyethyleneglycol 1000 succinate (MW=530). TPGS is anon-ionic surfactant (HLB=16-18) with the structure of Scheme II:

Various chemical derivatives of vitamin E TPGS including ester and etherlinkages of various chemical moieties are included within the definitionof vitamin E TPGS.

Polyethylene glycol: Polyethylene glycol (PEG) is a hydrophilic,polymerized form of ethylene glycol, consisting of repeating units ofthe chemical structure—(CH₂—CH₂—O—).

AUC: AUC is the area under the plasma concentration-time, commonly usedin pharmacokinetics to quantitate the percentage of drug absorption andelimination. A high AUC generally indicates that the drug willsuccessfully reach the target tissue or organ.

Poloxamers or Pluronics: are synthetic block copolymers of ethyleneoxide and propylene oxide having the general structure:

OH (OCH₂CH₂)a (OCH₂CH₂CH₂)b (OCH₂CH₂)a H

The following variants based on the values of a and b are commerciallyavailable from BASF Performance Chemicals (Parsippany, N.J.) under thetrade name Pluronic and which consist of the group of surfactantsdesignated by the CTFA name of Poloxamer 108, 188, 217, 237, 238, 288,338, 407, 101, 105, 122, 123, 124, 181, 182, 183, 184, 212, 231, 282,331, 401, 402, 185, 215, 234, 235, 284, 333, 334, 335, and 403. For themost commonly used poloxamers 124, 188, 237, 338 and 407 the values of aand b are 12/20, 79/28, 64137, 141/44 and 101/56, respectively.

Solutol HS-15: is a polyethylene glycol 660 hydroxystearate manufacturedby BASF (Parsippany, N.J.). Apart from free polyethylene glycol and itsmonoesters, di-esters are also detectable. According to themanufacturer, a typical lot of Solutol HS-15 contains approximately 30%free polyethylene glycol and 70% polyethylene glycol esters.

Other surfactants: Other surfactants useful in the invention includeascorbyl-6 palmitate (Roche Vitamins, Nutley N.J.), stearylamine, andsucrose fatty acid esters (Mitsubishi Chemicals). Custom surfactantsinclude those compounds with polar water-loving heads and hydrophobictails, such as a vitamin E derivative comprising a peptide bondedpolyglutamate attached to the ring hydroxyl and pegylated phytosterol.

Hydrophile-lipophile balance: An empirical formula used to indexsurfactants. Its value varies from 1-45 and in the case of non-ionicsurfactants from about 1-20. In general for lipophilic surfactants theHLB is less than 10 and for hydrophilic ones the HLB is greater than 10.

Biocompatible: Capable of performing functions within or upon a livingorganism in an acceptable manner, without undue toxicity orphysiological or pharmacological effects.

Substantially ethanol-free: A composition having an ethanolconcentration less than about 1.0% (w/v) ethanol.

Emulsion: A colloidal dispersion of two immiscible liquids in the formof droplets, whose diameter, in general, are between 0.1 and 3.0 micronsand which is typically optically opaque, unless the dispersed andcontinuous phases are refractive index matched. Such systems possess afinite stability, generally defined by the application or relevantreference system, which may be enhanced by the addition of amphiphilicmolecules or viscosity enhancers.

Microemulsion: A thermodynamically stable isotropically clear dispersionof two immiscible liquids, such as oil and water, stabilized by aninterfacial film of surfactant molecules. The microemulsion has a meandroplet diameter of less than 200 nm, in general between 10-50 nm. Inthe absence of water, mixtures of oil(s) and non-ionic surfactant(s)form clear and isotropic solutions that are known as self-emulsifyingdrug delivery systems (SEDDS) and have successfully been used to improvelipophilic drug dissolution and oral absorption

Aqueous Medium: A water-containing liquid which can containpharmaceutically acceptable additives such as acidifying, alkalizing,buffering, chelating, complexing and solubilizing agents, antioxidantsand antimicrobial preservatives, humectants, suspending and/or viscositymodifying agents, tonicity and wetting or other biocompatible materials.

Therapeutic Agent: Any compound natural of synthetic which has abiological activity, is soluble in the oil phase and has anoctanol-buffer partition coefficient (Log P) of at least 2 to ensurethat the therapeutic agent is preferentially dissolved in the oil phaserather than the aqueous phase. This includes peptides, non-peptides andnucleotides. Lipid conjugates/prodrugs of water soluble molecules arewithin the scope of therapeutic agent.

Chemotherapeutic: Any natural or synthetic molecule which is effectiveagainst one or more forms of cancer, and particularly those moleculeswhich are slightly or completely lipophilic or which can be modified tobe lipophilic. This definition includes molecules which by theirmechanism of action are cytotoxic (anti-cancer agents), those whichstimulate the immune system (immune stimulators) and modulators ofangiogenesis. The outcome in either case is the slowing of the growth ofcancer cells.

Chemotherapeutics include Taxol (paclitaxel) and related moleculescollectively termed taxoids, taxines or taxanes. The structure ofpaclitaxel is shown in the figure below (Scheme III).

Included within the definition of “taxoids”are various modifications andattachments to the basic ring structure (taxoid nucleus) as may be shownto be efficacious for reducing cancer cell growth and to partition intothe oil (lipid phase) and which can be constructed by organic chemicaltechniques known to those skilled in the art. The structure of thetaxoid nucleus is shown in Scheme IV.

Chemotherapeutics include podophyllotoxins and their derivatives andanalogues. The core ring structure of these molecules is shown in thefollowing figure (Scheme V):

Another important class of chemotherapeutics useful in this inventionare camptothecins, the basic ring structure of which is shown in thefollowing figure, but includes any derivatives and modifications to thisbasic structure which retain efficacy and preserve the lipophiliccharacter of the moleculeshown below (Scheme VI).

Another preferred class of chemotherapeutics useful in this inventionare the lipophilic anthracyclines, the basic ring structure of which isshown in the following figure (Scheme VII):

Suitable lipophilic modifications of Scheme VII include substitutions atthe ring hydroxyl group or sugar amino group.

Another important class of chemotherapeutics are compounds which arelipophilic or can be made lipophilic by molecular chemosyntheticmodifications well known to those skilled in the art, for example bycombinatorial chemistry and by molecular modelling, and are drawn fromthe following list: Taxotere, Amonafide, Illudin S,6-hydroxymethylacylfulvene Bryostatin 1, 26-succinylbryostatin 1,Palmitoyl Rhizoxin, DUP 941, Mitomycin B, Mitomycin C, Penclomedine.Interferon α2b, angiogenesis inhibitor compounds, Cisplatin hydrophobiccomplexes such as 2-hydrazino-4,5-dihydro-1H-imidazole with platinumchloride and 5-hydrazino-3,4-dihydro-2H-pyrrole with platinum chloride,vitamin A, vitamin E and its derivatives, particularly tocopherolsuccinate.

Other compounds useful in the invention include:1,3-bis(2-chloroethyl)-1-nitrosurea (“carmustine” or “BCNU”),5-fluorouracil, doxorubicin (“adriamycin”), epirubicin, aclarubicin,Bisantrene(bis(2-imidazolen-2-ylhydrazone)-9,10-anthracenedicarboxaldehyde,mitoxantrone, methotrexate, edatrexate, muramyl tripeptide, muramyldipeptide, lipopolysaccharides, 9-b-d-arabinofuranosyladenine(“vidarabine”) and its 2-fluoro derivative, resveratrol, retinoic acidand retinol, Carotenoids, and tamoxifen.

Other compounds useful in the application of this invention include:Palmitoyl Rhizoxin, DUP 941, Mitomycin B, Mitomycin C, Penclomedine,Interferon α2b, Decarbazine, Lonidamine, Piroxantrone, Anthrapyrazoles,Etoposide, Camptothecin, 9-aminocamptothecin, 9-nitrocamptothecin,camptothecin-11 (“Irinotecan”), Topotecan, Bleomycin, the Vincaalkaloids and their analogs [Vincristine, Vinorelbine, Vindesine,Vintripol, Vinxaltine, Ancitabine], 6-aminochrysene, and navelbine.

Other compounds useful in the application of the invention are mimeticsof taxol, eleutherobins, sarcodictyins, discodermolides andepothiolones.

Taking into account these definitions, the present invention is directedto pharmaceutical compositions in the form of emulsions, micellarsolutions or self-emulsifying drug delivery systems which aresubstantially free of ethanol solvent.

The therapeutic agents of the compositions of this invention caninitially be solublized in ethanol. However, the ethanol is removed inorder to form a substantially ethanol-free composition. The ethanolconcentration is less than 1% (w/v), preferably less than 0.5%, and mostpreferably less than 0.3%. The therapeutic agents can also besolubilized in methanol, propanol, chloroform, isopropanol, butanol andpentanol. These solvents are also removed prior to use.

The compositions of the invention contain α-tocopherol as a carrier fortherapeutic drugs, which can be administered to animals or humans viaintravascular, oral, intramuscular, cutaneous and subcutaneous routes.Specifically, the emulsions can be given by any of the following routes,among others: intraabdominal, intraarterial, intraarticular,intracapsular, intracervical, intracranial, intraductal, intradural,intralesional, intralocular, intralumbar, intramural, intraocular,intraoperative, intraparietal, intraperitoneal, intrapleural,intrapulmonary, intraspinal, intrathoracic, intratracheal,intratympanic, intrauterine, and intraventricular. The emulsions of thepresent invention can be nebulized using suitable aerosol propellantswhich are known in the art for pulmonary delivery of lipophiliccompounds.

In its first aspect, the invention is directed to the use ofα-tocopherol as the hydrophobic dispersed phase of emulsions containingwater insoluble, poorly water soluble therapeutic agents, water solubletherapeutic agents which have been modified to be less water soluble ormixtures thereof. Also called vitamin E, α-tocopherol is not a typicallipid oil. It has a higher polarity than most lipid oils, particularlytriglycerides, and is not saponifiable. It has practically no solubilityin water.

In the second aspect, the invention is a an α-tocopherol emulsion in theform of a self-emulsifying system where the system is to be used for theoral administration of water insoluble (or poorly water soluble or watersoluble agents modified to be less water soluble or mixtures thereof)drugs where that is desired. In this embodiment, an oil phase withsurfactant and drug or drug mixture is encapsulated into a soft or hardgelatin capsule. Suitable solidification agents with melting points inthe range of 40 to 60° C. such as high molecular weight polyethyleneglycols (MW>1000) and glycerides such as those available under the tradename Gellucires (Gattefose Corp. Saint Priest, France) can be added toallow filling of the formulation into a hard gelatin capsule at hightemperature. Semi-solid formulations are formed upon room temperatureequilbration. Upon dissolution of the gelatin in the stomach andduodenum, the oil is released and forms a fine emulsion with a meandroplet diameter of between 2-5 microns spontaneously. The emulsion isthen taken up by the microvilli of the intestine and released into thebloodstream.

In a third aspect, the invention comprises microemulsions containingα-tocopherol. Microemulsions refer to a sub-class of emulsions where theemulsion suspension is essentially clear and indefinitely stable byvirtue of the extremely small size of the oil/drug microaggregatesdispersed therein.

In a fourth aspect of the invention, PEGylated vitamin E (TPGS) is usedas a primary surfactant in emulsions of vitamin E. PEGylated vitamin Eis utilized as a primary surfactant, a stabilizer and also as asupplementary solvent in emulsions of vitamin E. Polyethylene glycol(PEG) is also useful as a secondary solvent in the emulsions of thisinvention.

The α-tocopherol concentration of the emulsions of this invention can befrom about 2 to about 10% w/v. The ratio of α-tocopherol to TPGS isoptimally from about 1:1 to about 10:1 (w/w).

The emulsions of the invention may further include surfactants such asascorbyl-6 palmitate; stearylamine; sucrose fatty acid esters andvarious vitamin E derivatives comprising α-tocopherol nicotinate,tocopherol phosphate, and nonionic, synthetic surfactant mixtures,containing a fluorine-containing surfactant, such as the Zonyl brandseries and a polyoxypropylene-polyoxyethylene glycol nonionic blockcopolymer.

The emulsions of the invention can comprise an aqueous medium. Theaqueous phase has an osmolality of approximately 300 mOsm and mayinclude potassium or sodium chloride sorbitol, mannitol, polyethyleneglycol, propylene glycol albumin, polypep and mixtures thereof. Thismedium can also contain various additives to assist in stabilizing theemulsion or in rendering the formulation biocompatible. Acceptableadditives include acidifying agents, alkalizing agents, antimicrobialpreservatives, antioxidants, buffering agents, chelating agents,suspending and/or viscosity-increasing agents, and tonicity agents.Preferably, agents to control the pH, tonicity, and increase viscosityare included. Optimally, a tonicity of at least 250 mOsm is achievedwith an agent which also increases viscosity, such as sorbitol orsucrose.

The emulsions of the invention for intravenous injection have a particlesize of 10 to 500 nm, preferably 10 to 200 nm and most preferably 10 to100 nm. For intravenous emulsions, the spleen and liver will eliminateparticles greater than 500 mn in size through the RES.

A preferred form of the invention includes paclitaxel, a verywater-insoluble cytotoxin used in the treatment of uterine cancer andother carcinomas. An emulsion composition of the present inventioncomprises a solution of vitamin E containing paclitaxel at aconcentration of up to 20 mg/mL, four times that currently available byprescription, and a biocompatible surfactant such that the emulsionmicrodroplets are less than 0.2 microns and are terminally sterilizableby filtration.

A further embodiment of the invention is a method of treating carcinomascomprising the parenteral administration of a bolus dose of paclitaxelin vitamin E emulsion with and without PEGylated vitamin E byintravenous injection once daily or every second day over a therapeuticcourse of several weeks. Such method can be used for the treatment ofcarcinomas of the breast, lung, skin and uterus.

The general principles of the present invention may be more fullyappreciated by reference to the following non-limiting examples.

EXAMPLES Example 1 Dissolution of Paclitaxel in α-Tocopherol

α-Tocopherol was obtained from Sigma Chemical Company (St Louis Mo.) inthe form of a synthetic dl-α-tocopherol of 95% purity prepared fromphytol. The oil was amber in color and very viscous. Paclitaxel waspurchased from Hauser Chemical Research (Boulder Colo.), and was 99.9%purity by HPLC. Paclitaxel 200 mg was dissolved in 6 mL of dry absoluteethanol (Spectrum Chemical Manufacturing Corp, Gardenia Calif.) andadded to 1 gm α-tocopherol. The ethanol was then removed by vacuum at42° C. until the residue was brought to constant weight. Independentstudies showed that the ethanol content was less than 0.3% (w/v).

The resultant solution was clear, amber and very viscous, with a nominalconcentration of 200 mg/gm (w/w) paclitaxel in α-tocopherol. Higherconcentrations of Paclitaxel (up to 400 mg/gm, w/w) can be solubilizedin α-tocopherol.

Example 2 Anionic Surfactant Used to Prepare α-Tocopherol Emulsions

Paclitaxel 2 gm in 10 gm of α-tocopherol, prepared as described inExample 1, was emulsified with ascorbyl palmitate as the triethanolaminesalt by the following method. A solution consisting of ascorbic acid 20mM was buffered to pH 6.8 with triethanolamine as the free base to from2×buffer. 50 mL of the 2×buffer was placed in a Waring blender. 0.5 gmof ascorbyl-6-palmitate (Roche Vitamins and Fine Chemicals, NutleyN.J.), an anionic surfactant, was added and the solution blended at highspeed for 2 min at 40° C. under argon. The α-tocopherol containingpaclitaxel was then added into the blender with the surfactant andbuffer. Mixing was continued under argon until a coarse, milky,pre-emulsion was obtained, approximately after 1 min at 40° C. Water forinjection was then added, bringing the final volume to 100 mL.

The pre-emulsion was transferred to the feed vessel of a MicrofluidizerModel 110Y (Microfluidics Inc, Newton Mass.). The unit was immersed in abath to maintain a process temperature of approximately 60° C. duringhomogenization, and was flushed with argon before use. After priming,the emulsion was passed through the homogenizer in continuous re-cyclefor 10 minutes at a pressure gradient of about 18 kpsi across theinteraction head. The flow rate was about 300 mL/min, indicating thatabout 25 passes through the homogenizer resulted.

The resultant paclitaxel emulsion in an α-tocopherol vehicle was bottledin amber vials under argon and stored with refrigeration at 7° C. and25° C. Samples were taken at discrete time intervals for particle sizingand chemical analysis.

Data taken with a Nicomp Model 370 Submicron Particle Sizer (ParticleSizing Systems Inc, Santa Barbara Calif.) showed that the emulsion had amean particle diameter of 280 nm.

Example 3 Use of PEGylated Vitamin E (TPGS)

A ternary phase diagram was constructed for α-tocopherol, PEGylatedvitamin E (TPGS, vitamin-E polyoxyethyleneglycol-1000-succinate,obtained from Eastman Chemical Co., Kingsport Tenn.), and water. TPGSwas first melted at 42° C. and mixed gravimetrically with α-tocopherolat various proportions from 1 to 100% TPGS, the balance beingα-tocopherol. Mixtures were miscible at all concentrations. Water wasthen added to each mixture in such a way that the final waterconcentration was increased stepwise from zero to 97.5%. At each step,observations were made of the phase behavior of the mixture. Asappropriate, mixing was performed by vortexing and sonication, and themixture was heated or centrifuged to assess its phase composition.

A broad area of biphasic o/w emulsions suitable for parenteraladministration was found at water concentrations above 80%. Theemulsions formed were milky white, free flowing liquids that containeddisperse α-tocopherol microparticles stabilized by non-ionic surfactant.Also in this area, microemulsions potentially suitable as drug carrierswere observed at TPGS to oil ratios above about 1:1. At lower watercontent, a broad area containing transparent gels (reverse emulsions)was noted. Separating the two areas (high and low water content) is anarea composed of opaque, soap-like liquid crystals.

Phase diagrams of α-tocopherol with surfactant combinations, for exampleTPGS with a nonionic, anionic or cationic co-surfactant (for exampleglutamyl stearate, ascorbyl palmitate or Pluronic F-68), or drug can beprepared in a similar manner.

Example 4 α-Tocopherol Emulsion for Intravenous Delivery of Paclitaxel

A formulation of the following composition was prepared:

Paclitaxel 1.0 gm % α-tocopherol 3.0 gm % TPGS 2.0 gm %Ascorbyl-6-Palmitate 0.25 gm % Sorbitol 5.0 gm % Triethanolamine to pH6.8 Water qs to 100 mL

The method of preparation was as follows: synthetic α-tocopherol (RocheVitamins, Nutley N.J.), paclitaxel (Hauser, Boulder Colo.), ascorbyl6-palmitate (Aldrich Chemical Co, Milwaukee Wis.) and TPGS weredissolved in 10 volumes of anhydrous undenatured, ethanol (SpectrumQuality Products, Gardenia Calif.) with heating to 40-45° C. The ethanolwas then drawn off with vacuum until no more than 0.3% remained byweight.

Pre-warmed aqueous solution containing a biocompatible osmolyte andbuffer were added with gentle mixing and a white milk formedimmediately. This mixture was further improved by gentle rotation for 10minutes with continuous warming at 40-45° C. This pre-mixture at aboutpH 7 was then further emulsified as described below.

The pre-mixture at 40-45° C. was homogenized in an Avestin C5homogenizer (Avestin, Ottawa Canada) at 26 Kpsi for 12 minutes at 44° C.The resultant mixture contained microparticles of α-tocopherol with amean size of about 200 nm. Further pH adjustment was made with analkaline 1 M solution of triethanolamine (Spectrum Quality Products).

In order to avoid gelation of the TPGS during the early stage ofemulsification, all operations were performed above 40° C. and care wastaken to avoid exposure of the solutions to cold air by covering allvessels containing the mixture. Secondly, less than 2% TPGS shouldgenerally be dissolved in α-tocopherol oil before pre-emulsification,the balance of the TPGS being first dissolved in the aqueous bufferbefore the pre-emulsion is prepared. The solution gels at concentrationsof TPGS higher than 2%.

Physical stability of the emulsion was then examined by placing multiplevials on storage at 4° C. and 25° C. Over several months, vials wereperiodically withdrawn for particle sizing. Mean particle size, asdetermined with the Nicomp Model 370 (Particle Sizing Systems, SantaBarbara Calif.), is shown for the two storage temperatures in FIG. 1.The particle size distribution was bi-modal.

Example 5 Chemical Stability of Paclitaxel in an α-Tocopherol Emulsion

After emulsification, the formulation of Example 4 was analyzed forpaclitaxel on a Phenosphere CN column (5 microns, 150×4.6 mm). Themobile phase consisted of a methanol/water gradient, with a flow rate of1.0 mL/min. A UV detector set at 230 nm was used to detect andquantitate paclitaxel. A single peak was detected (FIG. 2), which had aretention time and mass spectrogram consistent with native referencepaclitaxel obtained from Hauser Chemical (Boulder Colo.).

Chemical stability of the emulsion of example 4 was examined by HPLCduring storage. The data of FIG. 3 demonstrate that paclitaxel remainsstable in the emulsion for periods of at least 3 months, independent ofthe storage temperature. Taken together, the data of FIG. 2 and 3demonstrate successful retention of drug potency and emulsion stabilitywhen stored at 4° C. for a period of at least 3 months.

Example 6 Paclitaxel Emulsion Formulation QWA

An emulsion of paclitaxel 10 mg/ml for intravenous drug delivery, havingthe following composition, was prepared as described in Example 4.

Paclitaxel 1.0 gm % α-tocopherol 3.0 gm % TPGS 1.5 gm %Ascorbyl-6-Palmitate 0.25 gm % Sorbitol 4.0 gm % Triethanolamine to pH6.8 Water qs to 100 mL

Example 7 Paclitaxel Emulsion Formulation QWB

A second emulsion of paclitaxel 10 mg/ml for intravenous drug delivery,having the following composition, was prepared as described in Example4.

Paclitaxel 1.0 gm % α-tocopherol 3.0 gm % TPGS 1.5 gm % Solutol HS-151.0 gm % Sorbitol 4.0 gm % Triethanolamine to pH 6.8 Water qs to 100 mL

Solutol HS-15 is a product of BASF Corp, Mount Olive N.J.

Example 8 10 mg/mL Paclitaxel Emulsion Formulation QWC

A third emulsion formulation of paclitaxel 10 mg/ml was prepared asfollows using Poloxamer 407 (BASF Corp, Parsippany N.J.) as aco-surfactant.

Paclitaxel 1.0 gm % α-tocopherol 6.0 gm % TPGS 3.0 gm % Poloxamer 4071.0 gm % Sorbitol 4.0 gm % Triethanolamine to pH 6.8 Water for injectionqs to 100 mL

In this example, 1.0 gm Poloxamer 407 and 1.0 gm paclitaxel weredissolved in 6.0 gm α-tocopherol with ethanol 10 volumes and gentleheating. The ethanol was then removed under vacuum. Separately, anaqueous buffer was prepared by dissolving 3.0 gm TPGS and 4.0 gmsorbitol in a final volume of 90 mL water for injection. Both oil andwater solutions were warmed to 45° C. and mixed with sonication to makea pre-emulsion. A vacuum was used to remove excess air from thepre-emulsion before homogenization.

Homogenization was performed in an Avestin C5 as already described. Thepressure differential across the homogenization valve was 25 kpsi andthe temperature of the feed was 42-45° C. A chiller was used to ensurethat the product exiting the homogenizer did not exceed a temperature of50° C. Flow rates of 50 mL/min were obtained during homogenization.After about 20 passes in a recycling mode, the emulsion became moretranslucent. Homogenization was continued for 20 min. Samples werecollected and sealed in vials as described before. A fine α-tocopherolemulsion for intravenous delivery of paclitaxel was obtained. The meanparticle diameter of the emulsion was 77 nm. Following 0.22μ sterilefiltration through a 0.22 micron Durapore filter (Millipore Corp,Bedford Mass.), the emulsion was filled in vials and stored at 4° C.until used for intravenous injection.

Example 9 5 mg/mL Paclitaxel Emulsion Formulation QWC

An additional emulsion of paclitaxel was prepared as described inExample 8 but incorporating 5 instead of 10 mg/ml of the drug. Thecomposition of this emulsion is as follows:

Paclitaxel 0.5 gm % α-tocopherol 6.0 gm % TPGS 3.0 gm % Poloxamer 4071.0 gm % Sorbitol 4.0 gm % Triethanolamine to pH 6.8 Water for injectionqs to 100 mL

Following homogenization as described in example 8, a somewhattranslucent emulsion of α-tocopherol and paclitaxel with a mean particlediameter of 52 nm was obtained. Following sterile filtration through a0.22 micron Durapore filter (Millipore Corp, Bedford Mass.), theemulsion was filled in vials and stored at 4° C. until used forintravenous injection. Drug losses on filtration were less than 1%.

Example 10 Paclitaxel Emulsion Formulation QWD

A fifth emulsion of α-tocopherol for intravenous administration ofpaclitaxel was prepared as follows:

Paclitaxel 0.5 gm % α-tocopherol 6.0 gm % TPGS 3.0 gm % Poloxamer 4071.5 gm % Polyethyleneglycol 200 0.7 gm % Sorbitol 4.0 gm %Triethanolamine to pH 6.8 Water for injection qs to 100 mL

Synthetic α-tocopherol USP-FCC obtained from Roche Vitamins (Nutley,N.J.) was used in this formation. Polyethyleneglycol 200 (PEG-200) wasobtained from Sigma Chemical Co.

Following homogenization, a somewhat translucent emulsion with a meanparticle diameter of 60 nm was obtained. Following 0.22μ sterilefiltration through a 0.22 micron Durapore filter (Millipore Corp,Bedford Mass.), the emulsion was filled in vials and stored at 4° C.until used for intravenous injection. Drug losses on filtration wereless than 1%.

Example 11 Dissolution of Paclitaxel in TPGS and Preparation of MicellarSolutions

We observed good solubility of paclitaxel in TPGS, about 100 mg drug per1.0 gm of TPGS. Micellar solutions of TPGS containing paclitaxel wereprepared as follows. A stock solution of paclitaxel in TPGS was made upby dissolving 90 mg paclitaxel in 1.0 gm TPGS at 45° C. with ethanol,which was then removed under vacuum. Serial dilutions were then preparedby diluting the paclitaxel stock with additional TPGS to obtainpaclitaxel in TPGS at concentrations of 0.1, 1, 5, 10, 25, 50, 75 and 90mg/mL. Using fresh test tubes, 100 mg of each paclitaxel concentrationin TPGS was dissolved in 0.9 mL water. All test tubes were mixed byvortex and by sonication at 45° C. Clear micellar solutions in waterwere obtained corresponding to final paclitaxel concentrations of 0.01,0.1, 0.5, 1.0, 2.5, 5.0, 7.5 and 9.0 mg/mL.

A Nicomp Model 370 laser particle sizer (Particle Sizing Systems, SantaBarbara Calif.) was used to examine the solutions. Particle sizes on theorder of 10 nm were obtained, consistent with the presence of micellesof TPGS and paclitaxel.

Micellar solutions of paclitaxel in TPGS containing up to 2.5 mg/mLpaclitaxel were stable for at least 24 hr whereas those at 5.0, 7.5 and9.0 mg/mL were unstable and drug crystals formed rapidly andirreversibly. These observations imply that paclitaxel remainssolubilized only in the presence of an α-tocopherol-rich domain withinthe emulsion particles. Thus, an optimum ratio of α-tocopherol to TPGSis needed in order to produce emulsions in which higher concentrationsof paclitaxel can be stabilized.

When adjusted to the proper tonicity and pH, micellar solutions haveutility for slow IV drip administration of paclitaxel to cancerpatients, although the AUC is expected to be low.

The utility of TPGS in α-tocopherol emulsions is a synergy of severaldesirable characteristics. First, it has its own affinity forpaclitaxel, probably by virtue of the α-tocopherol that makes up thehydrophobic portion of its molecular structure. Secondly, interfacialtension of TPGS in water with α-tocopherol is about 10 dynes/cm,sufficient to emulsify free α-tocopherol, especially when used with aco-surfactant. Third, polyoxyethylated surfactants such as TPGS, havewell established, superior properties as a “stealth coat” for injectableparticles, by dramatically reducing trapping of the particles in theliver and spleen, as is well known in the art. But the unexpected andunique finding with TPGS as a surfactant for α-tocopherol emulsions, wasthe finding of all three desirable characteristics in a single molecule.An additional advantage of TPGS is the fact that it forms stableself-emulsifying systems in mixtures with oils and solvents such aspropylene glycol and polyethylene glycol, suggesting a synergy when usedwith α-tocopherol for oral drug delivery.

When adjusted to the proper tonicity and pH, micellar solutions haveutility for slow IV drip administration of paclitaxel to cancerpatients, although the AUC is expected to be low.

Example 12 20 mg/mL Paclitaxel Emulsion Formulation

A coarse, emulsion containing 20 mg/mL paclitaxel in α-tocopherol wasobtained with 5% α-tocopherol and 5% TPGS by the methods described inExample 4, simply by increasing the concentrations. No effort was madeto test higher concentrations simply because no further increase isnecessary for clinically useful intravenous emulsions.

Example 13 Use of Other PEG Surfactants in α-Tocopherol Emulsions

A variety of other pegylated surfactants, for example Triton X-100, PEG25 propylene glycol stearate, Brij 35 (Sigma Chemical Co), Myrj 45, 52and 100, Tween 80 (Spectrum Quality Products), PEG 25 glyceryl trioleate(Goldschmidt Chemical Corp, Hopewell Va.), have utility in emulsifyingα-tocopherol.

However, experiments with other pegylated surfactants failed toconvincingly stabilize paclitaxel in an α-tocopherol emulsion. Todemonstrate the unique utility of TPGS, three emulsions were prepared asdescribed in Example 9, but Tween 80 and Myrj 52 were substituted forTPGS as the primary surfactant in separate emulsions. These twosurfactants were chosen because Tween 80 and Myrj 52 have HLB valuesessentially equivalent to TPGS and make reasonably good emulsions ofα-tocopherol. However, when 5 mg/mL paclitaxel was included in theformulation, drug crystallization was noted very rapidly afterpreparation of the pre-emulsion, and the processed emulsions of Tween 80and Myrj 52 were characterized as coarse, containing rod-shapedparticles up to several microns in length, consistent with crystals ofpaclitaxel. Unlike the TPGS emulsion, which passed readily through a0.22 micron filter with less than 1% loss of drug, the Tween and Myijemulsions were unfilterable because of the presence of this crystallinedrug material.

There are several possible explanations for the unexpected improvementof the α-tocopherol paclitaxel emulsions with TPGS. The drug has goodsolubility in TPGS, up to about 100 mg/mL. Most likely it is thestrength of the affinity of paclitaxel benzyl side chains with theplanar structure of the α-tocopherol phenolic ring in the TPGS moleculethat stabilizes the complex of drug and carrier. In addition thesuccinate linker between the α-tocopherol and PEG tail is a novelfeature of this molecule that distinguishes its structure from otherPEGylated surfactants tested.

Example 14 Poloxamer-based α-Tocopherol Emulsion

α-tocopherol 6.0 gm % Poloxamer 407 2.5 gm % Ascorbyl Palmitate 0.3 gm %Sorbitol 6.0 gm % Triethanolamine to pH 7.4 Water qs to 100 mL

An α-tocopherol emulsion was prepared using Poloxamer 407 (BASF) as theprimary surfactant. The white milky pre-mixture was homogenized withcontinuous recycling for 10 minutes at 25 Kpsi in a C5 homogenizer(Avestin, Ottawa Canada) with a feed temperature of 45° C. and a chillerloop for the product out set at 15° C. A fine, sterile filterableemulsion of α-tocopherol microparticles resulted. However, when thisformulation was made with paclitaxel, precipitation of the paclitaxelwas noted following overnight storage in the refringerator, againunderlying the superior utility of TPGS as the principle surfactant.

Example 15 Lyophilized Emulsion Formulation

Maltrin M100 (Grain Processing Corporation, Muscatine Iowa) was added asa 2×stock in water to the emulsion of Example 14. Aliquots were thenfrozen in a shell freezer and lyophilized under vacuum. Onreconstitution with water, a fine emulsion was recovered.

Lyophilized formulations have utility where the indefinite shelf life ofa lyophilized preparation is preferred. Lyophilizable formulationscontaining other saccharides, such as mannitol, albumin or PolyPep fromSigma Chemicals, St. Louis, Mo. can also be prepared.

Example 16 In Vitro Release of Paclitaxel from α-Tocopherol Emulsions

One of the desired characteristics of a drug delivery vehicle is toprovide sustained release of the incorporated drug, a characteristicquite often correlated with improved pharmacokinetics and efficacy. Inparticular, long-circulating emulsions of paclitaxel can improve thedelivery of the drug to cancer sites in the body. We have surprisinglyfound that the emulsions of the present invention do provide sustainedrelease of paclitaxel when compared to the only FDA-approved formulationof paclitaxel at this time [Taxol®, Bristol Myers Squibb(BMS), PrincetonN.J.]. Emulsions were prepared having paclitaxel concentrations of 6mg/mL (QWA) and 7 mg/mL (QWB). For comparison, Taxol contains 6 mg/ml ofpaclitaxel dissolved in ethanol:cremophore EL 1:1 (v/v). In vitrorelease of paclitaxel from the different formulations into a solution ofphosphate-buffered saline (PBS) at 37° C. was monitored using a dialysismembrane that is freely permeable to paclitaxel (MW cut-off of 10kilodaltons). Quantification of the drug in pre- and post-dialysissamples was performed by HPLC. Drug release profiles in terms of bothpercent release and concentration of paclitaxel released over time weregenerated. As can be seen from the data in FIG. 4, less than 5% ofpaclitaxel was dialyzed from the emulsions over 24 hr, whereas about 12%was recovered outside the dialysis bag from the marketed BMSformulation. This indicates that drug release from the emulsion wassignificantly slowed relative to the commercially available solution.

Example 17 Biocompatibility of α-Tocopherol Emulsions ContainingPaclitaxel

An acute single-dose toxicity study was performed. Mice 20-25 gm eachwere purchased and acclimatized in an approved animal facility. Groupsof mice (n=3) received doses of the formulation containing from 30 to 90mg/kg paclitaxel in the α-tocopherol emulsion prepared as described inExample 6. All injections were given intravenously by tailvein bolus.

Although all injections were given by bolus IV push, no deaths orimmediate toxicity were observed at any dose, even at 90 mg/kg. Theresults for body weight are shown in Table 1. Weight loss was 17% in thehighest group but all groups, even at 90 mg/kg, recovered or gained bodyweight over a period of 10 days post injection.

A vehicle toxicity study was also done. Animals receiving drug-freeemulsion grew rapidly, and gained slightly more weight than animalsreceiving saline or not injected. This was attributed to the vitamin andcalorie content of the formulation.

We observed a maximal tolerable dose (MTD) for paclitaxel of greaterthan 90 mg/kg (Table 1), with no adverse reactions noted. This is morethan double the best literature values reported, in which deaths wereobserved at much smaller doses. Taxol, the FDA-approved BMS formulation,causes death in mice at bolus intravenous doses of 10 mg/kg, a findingrepeated in our hands. In the rat, BMS Taxol was uniformly fatal at alldilutions and dose regimes we tested. In contrast, the composition ofExample 6 was well tolerated in rats, and is even improved overTaxotere, a less toxic paclitaxel analogue commercially marketed byRhone-Poulenc Rorer.

One possible explanation for the high drug tolerance is that theemulsion is behaving as a slow-release depot for the drug as suggestedfrom the in vitro release data in Example 16.

TABLE 1 Average Body Weight Change of Mice Treated with PaclitaxelEmulsion Treatment Number of BW Change (gm) (dose, mg/kg) Animals Day 2Day 7 Saline 4 1.0 3.4 Vehicle 4 1.2 3.5 Paclitaxel Emulsion 2 −1.0 2.2(QWA) (36.3) Paclitaxel Emulsion 4 −1.8 1.7 (QWA) (54.4) PaclitaxelEmulsion 4 −1.5 1.6 (QWA) (72.6) Paclitaxel Emulsion 1 −1.6 (QWA) (90.7)

Example 18 Efficacy Evaluation of Paclitaxel Emulsion

The paclitaxel emulsion of Example 6 was also evaluated for efficacyagainst staged B16 melanoma tumors in nude mice and the data is shown inTable 2. Once again, the marketed product BMS Taxol was used as areference formulation. Tumor cells were administered subcutaneously andtherapy started by a tail vein injection at day 4 post-tumoradministration at the indicated dosing schedule. Efficacy was expressedas percent increase in life-span (% ILS).

The following conclusions can be drawn from the data in Table 2: a) anincreased life span of about 10% was obtained by administration of BMSTaxol at 10 mg/kg Q2Dx4, b) %ILS values improved to 30-50% byadministration of the α-tocopherol emulsion of paclitaxel at 30, 40 or50 mg/kg Q2Dx4, dose levels made possible by the higher MTD, c) a nicedose response was observed when the emulsion was administered at 30, 50and 70 mg/kg Q4Dx3, with about 80 % ILS being observed at 70 mg/kg and,d) even at 90 mg/kg dosed only once at day 4, there was about 36 % ILS.These data clearly illustrate the potential of the emulsions of thepresent invention to substantially improve the efficacy of paclitaxel.

Example 19 Efficacy Evaluation of Paclitaxel Emulsions

The emulsions of examples 6, 7 and 8 (QWA, QWB and QWC respectively)were compared for efficacy against B16 melanoma in mice; BMS taxol wasagain used as a reference formulation. Methods essentially identical tothose of Example 18 were used. The data from this study is summarized inTable 3. Efficacy was expressed as: a) percent tumor growth inhibition(% T/C, where T and C stand for treated and control animals,respectively); b) tumor growth delay value (T-C), and c) log cell killwhich is defined as the ratio of the T-C value over 3.32 33 tumordoubling time. The latter parameter for this particular tumor model wascalculated to be 1.75 days. As can be seen from the results in Table 3,all measures of efficacy: tumor growth inhibition, tumor growth delayvalue and log cell kill demonstrate superior efficacy of α-tocopherolemulsions as a drug delivery vehicle over BMS Taxol, particularly whenthe emulsions were dosed every four days at 70 mg/kg. As explained inExample 16, this increased efficacy is likely a result of improved drugbiocompatibility and/or sustained release.

TABLE 2 Survival of Mice with B16 Tumors Treated with QWA and BMS TaxolMean Survival Treatment Group & Time, Days % ILS^(b) (vs vehicle)Schedule (Mean ± S.E.M^(a)) (Mean ± S.E.M) Vehicle Control 13.2 ± 0.9 —(Days 4, 8, 12) Saline Control 15.8 ± 1.2 19.7 ± 8.6  (Days 4, 8, 12)BMS Taxol (10 mg/kg) 16.4 ± 0.7 24.2 ± 5.4  (Days 4, 6, 8, 10) QWA (30mg/kg) 19.2 ± 1.4 45.4 ± 10.3 (Days 4, 6, 8) QWA (40 mg/kg) 21.3 ± 1.461.4 ± 10.3 (Days 4, 6, 8) QWA (50 mg/kg) 18.8 ± 0.7 42.4 ± 5.7  (Days4, 6, 8) QWA (30 mg/kg) 15.3 ± 0.8 15.9 ± 6.4  (Days 4, 8, 12) QWA (50mg/kg) 20.7 ± 1.3 56.8 ± 9.5  (Days 4, 8, 12) QWA (70 mg/kg) 24.2 ± 0.983.3 ± 6.4  (Days 4, 8, 12) QWA (90 mg/kg) 18.0 ± 0.6 36.4 ± 4.4  (Day 4only) ^(a)SEM = Standard Error of Mean ^(b)% ILS = % Increase inLifespan = [(T − C)/C] × 100 where: T = mean survival of treated C =mean survival of control according to the NCI standards an ILS valuegreater than 50% indicates significant anti-tumor activity.

TABLE 3 Comparison of 3 paclitaxel emulsions and BMS taxol againstearly-stage B16 melanoma Total Median tumor Median tumor wt. Test DosageDosing Schedule Dose wt. on day 15 on day 18 mg % T/C T-C Log cellArticle mg/kg/day (days) (mg/kg) (mg) (range) Day 15 (days) kill totalControl 0 4, 6, 8, 10 0 836 2139 — — — BMS Taxol 20 4, 6, 8, 10 80 3831217 46 2 0.34 QWA 20 4, 6, 8, 10 80 381 1197 46 2 0.34 QWA 40 4, 6, 8,10 160 104 306 12 7 1.2 QWA 70 4, 8, 12, 16, 20 350 15 11 ˜2 QWB 20 4,6, 8, 10 80 197 653 24 5 0.86 QWB 30 4, 6, 8, 10 120 139 449 17 5 0.86QWB 40 Toxic QWC 20 4, 6, 8, 10 80 319 848 34 3 0.52 QWC 40 4, 6, 8, 10160 53 194 6 8 1.4 QWC 70 4, 8, 12, 16, 20 350 33 66 4 >15 >2.6 TumorDoubling Time calculated to be 1.75 days. % T/C = Tumor GrowthInhibition (Day 15) = (median tumor wt. of treated/median tumor wt.control) × 100 T-C = Tumor Growth Delay value = median time fortreatment group (T) and control group (C) tumors to reach apredetermined size (usually 750-1000 mg) Log cell kill = (T-Cvalue)/(3.32 × tumor doubling time)

Example 20 Seff-emulsification of an α-Tocopherol/Tagat TO Mixture

α-tocopherol 2.0 gm and Tagat TO (Goldschmidt Chemical Corp, HopewellVa.) 800 mg were dissolved together. About 80 mg of the oily mixture wastransferred to a test tube and water was then added. With gentle handmixing, there was immediate development of a rich milky emulsion,consistent with “self-emulsifying systems” proposed as drug deliverysystems, in which surfactant-oil mixtures spontaneously form an emulsionupon exposure to aqueous media.

Example 21 Self-emulsifying Formulation Containing Paclitaxel

Paclitaxel 50 mg/ml was prepared in α-tocopherol by the method describedin Example 1. Tagat TO 20% (w/w) was added. The resultant mixture wasclear, viscous and amber in color. A 100 mg quantity of the oily mixturewas transferred to a test tube. On addition of 1 mL of water, withvortex mixing, a fine emulsion resulted.

Example 22 Self-emulsifying Formulation of Paclitaxel

Paclitaxel 50 mg/ml was prepared in α-tocopherol by the method describedin Example 1. After removal of the ethanol under vacuum, 20% TPGS and10% polyoxyethyleneglycol 200 (Sigma Chemical Co) were added by weight.A demonstration of the self-emulsification ability of this system wasthen performed by adding 20 mL of deionized water to 100 mg of the oilymixture at 37° C. Upon gentle mixing, a white, thin emulsion formed,consisting of fine emulsion particles demonstrated with the MalvernMastersizer (Malvern Instruments, Worcester Mass.) to have a mean sizeof 2 microns, and a cumulative distribution 90% of which was less than10 microns.

Example 23 Etoposide Emulsion Formulation in α-Tocopherol

Etoposide 4 mg (Sigma Chemical Co) was dissolved in the followingsurfactant-oil mixture:

Etoposide 4 mg α-tocopherol 300 mg TPGS 50 mg Poloxamer 407 50 mg

Ethanol and gentle warming was used to form a clear amber solution ofdrug in oil. The ethanol was then removed under vacuum.

A pre-emulsion was formed by adding 4.5 mL of water containing 4%sorbitol and 100 mg TPGS at 45° C. with sonication. The particle sizewas further reduced by processing in an Emulsiflex 1000 (Avestin, OttawaCanada). The body of the Emulsiflex 1000 was fitted with a pair of 5 mLsyringes and the entire apparatus heated to 45° C. before use. The 5 mLof emulsion was then passed through it by hand approximately 10 times. Afree flowing, practical emulsion of etoposide in an α-tocopherol vehicleresulted.

We note that the solubilized form of etopside in α-tocopherol can alsobe used as an oral dosage form by adaption of the methods of thepreceding examples.

Example 24 Dissolution of Ibuprofen or Griseofulvin in α-Tocopherol

Ibuprofen is a pain-killer, and may be administered by injection whenrequired if there is danger that the drug will irritate the stomach. Thefollowing solution of ibuprofen in α-tocopherol may be emulsified forintravenous administration.

Ibuprofen (Sigma Chemicals), 12 mg. crystalline, dissolved withoutsolvent in α-tocopherol, 120 mg, by gentle heating. The resultant 10%solution of ibuprofen in vitamin E can be emulsified by the methodsdescribed in Examples 4, 6, 7, 8 or 22.

An antifungal compound, griseofulvin, 12 mg, was first dissolved in 3 mLof anhydrous ethanol; α-tocopherol was then added, 180 mg, and theethanol was removed with gentle heating under vacuum. The resultantsolution of griseofulvin in α-tocopherol is clear and can be emulsifiedby the methods described in Examples 4, 6, 7, 8 or 22.

Example 25 Vitamin E Succinate Emulsion Formulation

Vitamin E succinate has been suggested as a therapeutic for thetreatment of lymphomas and leukemias and for the chemoprevention ofcancer. The following is a composition and method for the emulsificationof vitamin E succinate in α-tocopherol. Sucrose ester S1170 is a productof Mitsubishi Kagaku Foods Corp, Tokyo Japan. Vitamin E succinate, asthe free acid, was obtained as a whitish powder from ICN Biomedicals,Aurora, OH. Emulsions incorporating other surfactants such as pluronics,and TPGS along with α-tocopherol and α-tocopherol succinate can beprepared in a similar manner with and without a therapeutic agent.

α-Tocopherol 8 gm and vitamin E succinate 0.8 gm were dissolved togetherin ethanol in a round bottom flask. After removal of the solvent, 100 mLof an aqueous buffer was added. The alkaline buffer consisting of 2%glycerol, 10 mM triethanolamine, and 0.5 gm % sucrose ester S1170. Aftermixing for 2 min, the pre-emulsion was transferred to an Avestin ModelC-5 homogenizer and homogenization was continued for about 12 minutes ata process feed temperature of 58° C. The pressure differential acrossthe interaction head was 25 to 26 kpsi. During homogenization, pH wascarefully monitored, and adjusted as required to pH 7.0. Care was takento exclude oxygen during the process. A fine white emulsion resulted.

Example 26 α-Tocopherol Levels in Esters

Levels of α-tocopherol in commerically available esters:tocopherol-acetate, -succinate, -nicotinate, -phosphate and TPGS wereeither provided by the vendor or determined by HPLC. The concentrationof free α-tocopherol in these solutions is less than 1.0%, generallyless than 0.5%.

Example 27 Resveratrol Emulsion Formulation

Resveratrol is a cancer chemopreventative first discovered as an extractof grape skins. It has been proposed as a dietary supplement.

Resveratrol was obtained from Sigma Chemical Co. While it dissolvedpoorly in ethanol, upon addition of 10 mg resveratrol, 100 mg ofα-tocopherol, 100 mg TPGS and ethanol, a clear solution formed rapidly.Upon removal of the ethanol, a clear amber oil remained.

The oily solution of resveratrol can be formulated as a self-emulsifyingsystem for oral delivery by the various methods of the precedingexamples.

Example 28 Muramyl Dipeptide Formulation

Muramyl dipeptides are derived from mycobacteria and are potentimmunostimulants representative of the class of muramyl peptides,mycolic acid and lipopolysaccharides. They have use, for example, in thetreatment of cancer, by stimulating the immune system to target andremove the cancer. More recently, muroctasin, a synthetic analog, hasbeen proposed to reduce non-specific side effects of the bacterial wallextracts.

N-acetylmuramyl-6-O-steroyl-1-alanyl-d-isoglutamine was purchased fromSigma Chemical Co. and 10 mg was dissolved in 100 mg α-tocopherol and 80mg TPGS. Ethanol was used as a co-solvent to aid in dissolution of thedipeptide, but was removed by evaporation under vacuum, leaving a clearsolution in α-tocopherol and surfactant.

This oil solution of the drug can be emulsified for parenteraladministration by the various methods of the preceding examples.

Example 29 Alcohol-containing Emulsion

In attempting to adapt the teachings of PCT WO 95/11039 to the oraladministration of paclitaxel, the following formulation was made.

Paclitaxel 0.125 gm α-tocopherol 0.325 gm TPGS 0.425 gm Ethanol 0.125 gm

As before, paclitaxel was dissolved in a α-tocopherol and TPGS withethanol, which was then removed under vacuum. By dry weight, residualethanol was less than 3 mg (0.3% w/w). Fresh anhydrous ethanol 0.125 gmwas then added back to the formulation. After mixing the suitability ofthe formulation for oral administration, as in a gelatin capsule, wassimulated by the following experiment. An aliquot of 100 mg of thefree-flowing oil was added to 20 mL of water at 37° C. and mixed gentlywith a vortex mixer. A fine emulsion resulted. But after twenty minutes,microscopy revealed the growth on large numbers of crystals in rosettes,characteristic of paclitaxel precipitation. It was concluded that thisformulation was not suitable for oral administration of paclitaxelbecause large amounts of the drug would be in the form of crystals onentry into the duodenum, where it would be prevented from uptake becauseof its physical form. We speculate that the excess of ethanol, incombination with the high ratio of TPGS to α-tocopherol, is responsiblefor the observed crystallization of the drug from this formulation.

Example 30 Alcohol-containing α-Tocopherol Emulsion

In attempting to adapt the teachings of PCT WO 95/11039 to theintravenous administration of paclitaxel, the following formulation wasmade:

Paclitaxel 0.050 gm α-tocopherol 0.100 gm Lecithin 0.200 gm Ethanol0.100 gm Butanol 0.500 gm

As before, paclitaxel was dissolved in α-tocopherol and TPGS withethanol, which was then removed under vacuum. By dry weight, residualethanol was less than 2 mg (0.5% w/w). Fresh anhydrous ethanol 0.100 gmand n-butanol 0.500 gm was then added back to the formulation. A clearoil resulted. The injection concentrate was tested for biocompatibilityin administration by standard pharmaceutical practice of admixture andsaline. About 200 mg of the oil was dropped into 20 mL of saline andmixed. Large flakes of insoluble material developed immediately and thegreatest amount of material formed dense deposits on the walls of thetest tube. The mixture was clearly unsuitable for parenteraladministration by any route, and we speculate that this is so regardlessof the identity of the drug contained in the formulation. By trial anderror we have learned that lecithin is a poor choice as surfactant forα-tocopherol by virtue of its low HLB (around 4). Other successfulexamples described here for fine emulsions suitable for parenteraladministration were all made with high HLB surfactants. Thesesurfactants include TPGS (HLB around 17), Poloxamer 407 (HLB about 22)and Tagat TO (HLB about 14.0). In general, we found that α-tocopherolemulsification is best performed with principal surfactants of HLB>10,preferably greater than 12. Lecithin is not in this class, although itcould be used as a co-surfactant. In comparison, typical o/w emulsionsof triglycerides are made with surfactants of HLB between 7 and 12,demonstrating that α-tocopherol emulsions are a unique class by virtueof the polarity and extreme hydrophobicity of the α-tocopherol, factorsthat also favor the solubility of lipophilic and slightly polarlipophilic drugs in α-tocopherol. See Emulsions: Theorv and Practice,2nd Ed. p.248 (1985).

We claim:
 1. A pharmaceutical composition, comprising a chemotherapeuticagent, wherein the chemotherapeutic agent is at least one of a taxoid, ataxane, or a taxine; a tocopherol; tocopherol polyethylene glycolsuccinate; polyethylene glycol; a surfactant; and an aqueous phase;wherein the composition is an emulsion or a microemulsion having an oilphase and a water phase, and wherein all of the chemotherapeutic agentis in the oil phase.
 2. The composition of claim 1, wherein the ratio ofthe tocopherol to the tocopherol polyethylene glycol succinate is fromabout 1:1 to about 10:1 w/w.
 3. The composition of claim 1, wherein saidsurfactant has an HLB of at least
 10. 4. The composition of claim 1,wherein said surfactant is at least one of an anionic, a cationic, anonionic, or a zwitterionic surfactant.
 5. The composition of claim 1,wherein said surfactant is at least one of apolyoxypropylene-polyoxyethylene glycol nonionic block co-polymer,ascorbyl-6-palmitate, stearylamine, or a sucrose fatty ester.
 6. Thecomposition of claim 5, wherein said surfactant is ascorbyl-6-palmitate.7. The composition of claim 1, wherein said chemotherapeutic agent ispaclitaxel.
 8. The composition of claim 1, wherein the emulsion has aparticle size of about 10 to about 500 nm.
 9. The composition of claim1, wherein said polyethylene glycol has an average molecular weight ofabout 100 to about
 600. 10. The composition of claim 9, wherein saidpolyethylene glycol has an average molecular weight of about
 400. 11.The composition of claim 8, wherein said particle size is from about 10to about 200 nm.
 12. The composition of claim 1, wherein said surfactantis a polyoxypropylene-polyoxyethylene glycol nonionic block co-polymer.13. The composition of claim 1, wherein said chemotherapeutic agent ispaclitaxel and said surfactant is a polyoxypropylene-polyoxyethyleneglycol nonionic block co-polymer.
 14. The composition of claim 12,wherein said block co-polymer has the structure:H(OCH₂CH₂)_(a)(OCH₂CH(CH₃))_(b)(OCH₂CH₂)_(a)OH wherein a is 101 and b is56.